Molecular mechanisms of excessive alcohol consumption and relapse-like behavior

过量饮酒行为和复发的行为和分子机制尚不完全清楚，对于绘制酒精中毒/依赖的病理过程至关重要。最近的证据表明，长期饮酒会导致大脑边缘关键区域兴奋性突触的加强，这些区域负责调节奖赏和饮酒行为。突触的这种强化高度依赖于alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic酸受体(AMPAR)的作用和突触结合；AMPAR是由兴奋性神经递质谷氨酸激活的快速作用离子通道受体。然而，增强的AMPAR信号在酒精强化和酒精寻求行为中的作用机制尚不清楚。因此，本申请中提出的实验试图通过一个高酒精消耗的临床前模型，酒精偏好(P-)大鼠，来阐明AMPAR介导的操作性酒精自我给药增加和增强复发样行为的行为和分子机制。初步数据表明，预先给予阿尼拉西坦(AMPAR的正变构调节剂)可增强AMPAR信号，增加可操作酒精的自我给药，并增强线索诱导的P-大鼠对酒精寻求的恢复，提示AMPAR活性的增强可能在促进饮酒增加和复发易感性中起关键作用。实验将进一步表征增强的AMPAR信号在调节操作者自我管理和复发样行为中的作用。AMPAR活性可以通过翻译后修饰(例如，AMPAR GluR1亚单位氨基酸残基831的磷酸化；pGluR1{831})来增强。利用免疫组织化学技术，实验将绘制边缘脑区pGluR1{831}亚单位在酒精自我给药或在线索诱导恢复期间暴露于与酒精相关的线索后的神经适应性变化，以确定可能影响AMPAR介导的增强的自我给药和酒精寻求增加的大脑区域。我们预测将在伏隔核(自我给药研究)或杏仁核(恢复研究)看到pGluR1{831}的变化，这些区域将被定位于研究AMPAR活性增强的神经解剖学控制--介导酒精自我给药和使用阿尼拉西坦寻求行为的便利化。对照实验将解决神经解剖学和增强剂的特异性以及非特异性运动效应。最后，我们将确定阿尼拉西坦诱导的酒精自我给药和寻找行为的增加是否依赖于钙/钙调蛋白依赖的蛋白激酶II(CaMKII；已知可以磷酸化GluR1并增强AMPAR活性)，在自我给药和使用KN93(CaMKII抑制剂)的恢复过程中阻断阿尼拉西坦诱导的作用。这些研究的关键发现将为AMPAR在过度饮酒行为和复发易感性中的相关机制提供新的见解。

The behavioral and molecular mechanisms underlying excessive alcohol drinking behavior and relapse are not fully understood and are vital for mapping the pathological course of alcoholism/dependence. Recent evidence indicates that chronic alcohol consumption leads to strengthening of excitatory synapses in key limbic brain regions that mediate reward and drinking behavior. This strengthening of synapses is highly dependent on the actions and synaptic incorporation of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs); fast action ion channel receptors that are activated by the excitatory neurotransmitter glutamate. However, the mechanistic role of enhanced AMPAR signaling in alcohol reinforcement and alcohol-seeking behavior remains unclear. Thus, the proposed experiments in this application seek to elucidate the behavioral and molecular mechanisms that underlie AMPAR-mediated increases in operant alcohol-self-administration and potentiated relapse-like behavior using a preclinical model of high alcohol consumption, the alcoholpreferring (P-) rat. Preliminary data indicate that enhancement of AMPAR signaling by pretreatment with aniracetam (positive allosteric modulator of AMPARs), increases operant alcohol self-administration and potentiates cue-induced reinstatement to alcohol seeking in P-rats, suggesting that enhanced AMPAR activity may be critical in facilitating increased drinking and susceptibility to relapse. Experiments will further characterize the role of enhanced AMPAR signaling in modulating operant self-administration and relapse-like behavior. AMPAR activity can be potentiated by post-translational modification (e.g. phosphorylation of the AMPAR GluR1 subunit amino acid residue 831;pGluR1{831}). Using immunohistochemistry techniques, experiments will map neuroadaptive changes in pGluR1{831} subunits in limbic brain regions after a history of alcohol self-administration or exposure to an alcohol-related cue during cue-induced reinstatement to determine brain regions that may influence enhanced AMPAR-mediated increases in self-administration and alcohol-seeking. We predict to see changes in pGluR1{831} in the nucleus accumbens (self-administration studies) or amygdala (reinstatement studies), and these regions will be targeted to investigate functional neuroanatomical control of enhanced AMPAR activity-mediated facilitation of alcohol self-administration and seeking behavior using aniracetam. Control experiments will address neuroanatomical and reinforcer specificity and non-specific locomotor effects. Lastly we will determine if aniracetam-induced increases in alcohol self-administration and seeking behavior are dependent on Ca2+/calmodulin-dependent protein kinase II (CamKII; known to phosphorylate GluR1 and enhance AMPAR activity) by blocking aniracetam-induced effects during self administration and reinstatement sessions with KN93 (CamKII inhibitor). Key findings from these studies will provide novel insight into AMPAR-related mechanisms in excessive alcohol drinking behavior and vulnerability to relapse.